Phosphatase-Mediated Hydrolysis of Linear Polyphosphates.

Polyphosphates are a group of phosphorus (P) containing molecules that are produced by a wide range of microorganisms and human activities. Although polyphosphates are ubiquitous in aquatic environments and are of environmental significance, little is known about their transformation and cycling. This study characterized the polyphopshate-hydrolysis mechanisms of several representative phosphatase enzymes and evaluated the effects of polyphosphate chain length, light condition, and calcium (Ca2+). 31P nuclear magnetic resonance (NMR) spectroscopy was used to monitor the dynamic changes of P molecular configuration during polyphosphate hydrolysis and suggested a terminal-only degradation pathway by the enzymes. Such mechanism enabled the quantification of the hydrolysis rates by measuring orthophosphate production over time. At the same initial concentration of polyphosphate molecules, the hydrolysis rates were independent of chain length. The hydrolysis of polyphosphate was also unaffected by light condition, but was reduced by the presence of Ca2+. The released orthophosphates formed Ca-phosphate precipitates in the presence of Ca2+, likely in amorphous phases. Results from this study lay the foundation for better understanding the chemical processes governing polyphosphate transport and transformation in various environmental settings.

Tumor accumulation of liposomal doxorubicin in three murine models: Optimizing delivery efficiency.

Systemic drug delivery to a solid tumor involves a sequence of steps that determine efficacy and survival. Extravasation from circulation at the tumor site is a critical step in this sequence since it regulates how much of the drug accumulates in the tumor. Despite its importance in determining outcomes, extravasation from circulation remains a "black box." The objective of this study is to develop predictive tools for optimization of drug delivery systems. By comparing pharmacokinetics of liposomal doxorubicin in tumor-free and tumor bearing mice we quantitatively assess the rate constants for distribution, elimination, and tumor accumulation. We then relate these rate constants to the tumor-type and drug delivery system. We compare tumor accumulation in three tumor types and show a 10-fold difference between a colorectal adenocarcinoma and a pancreatic adenocarcinoma. Finally, we show how quantitative predictions of changes in tumor accumulation can be used to optimize drug delivery systems.

Leakage kinetics of the liposomal chemotherapeutic agent Doxil: The role of dissolution, protonation, and passive transport, and implications for mechanism of action.

Doxil, a liposomal formulation of the chemotherapeutic drug doxorubicin, is FDA-approved for multiple indications. Doxil liposomes are designed to retain doxorubicin in circulation, minimize clearance by the mononuclear phagocyte system, and limit uptake in healthy tissue. Although pharmacokinetic data and survival statistics from clinical trials provide insight into distribution and efficacy, many details of the mechanism of action remain unresolved, despite the importance in translating liposome-based drug delivery systems to other molecules and cargo. Therefore, the objective of this study is to quantitatively assess the kinetics of doxorubicin leakage from Doxil liposomes. In contrast to previous studies, we consider three processes: dissolution of solid doxorubicin, protonation/deprotonation of soluble doxorubicin, and passive transport of neutral doxorubicin across the lipid bilayer of the liposomes. Experiments were performed for Doxil, Doxil-like liposomes, and Doxil-like liposomes with reduced cholesterol and pegylation. To mimic physiological conditions, we also performed experiments in serum and under slightly acidic conditions at pH5. We show that crystalline doxorubicin dissolution can be described by a first order rate constant of 1.0×10-9cms-1 at 37°C. Doxorubicin leakage can be described by first order rate constant for transport across the lipid bilayer with values in the range from 1 to 3×10-12cms-1 at 37°C. Based on these results we discuss implications for the mechanism of action, taking Doxil pharmacokinetics into account.

Brain microvascular endothelial cells resist elongation due to curvature and shear stress.

The highly specialized endothelial cells in brain capillaries are a key component of the blood-brain barrier, forming a network of tight junctions that almost completely block paracellular transport. In contrast to vascular endothelial cells in other organs, we show that brain microvascular endothelial cells resist elongation in response to curvature and shear stress. Since the tight junction network is defined by endothelial cell morphology, these results suggest that there may be an evolutionary advantage to resisting elongation by minimizing the total length of cell-cell junctions per unit length of vessel.